The machinery responsible for making proteins (e.g. ribosomal RNA, ribosomal proteins, translation factors, and tRNAs) is central to growth and development of all organisms. The regulation of the synthesis of the translation machinery in response to the nutritional state of the cell has been a central issue in the study of microbial physiology for at least forty years. More recently, it has also become clear that an understanding of the mechanisms responsible for rRNA transcription in Escherichia coli can provide fundamental insights into bacterial transcription in general. We are now poised to address questions central not only to our understanding of general transcription mechanisms, but also to how different regulatory systems work together. The questions addressed in the proposal are divided into three specific aims in arbitrary order: In the first aim, we propose to determine how a recognition element in bacterial promoters, the UP element, interacts with the RNA polymerase (RNAP) alpha subunit and whether it affects steps after recruitment of the enzyme to the complex, perhaps through alpha-sigma subunit interactions. In the second aim, we propose to determine how the rrn transcription factor FIS increases transcription of many promoters and how it can affect their responses to nutritional conditions. We propose to define the interactions between FIS and alpha at two rRNA promoters, rrnB P1 and at rrnE P1. In the third aim, we propose to determine the molecular details of regulation of rRNA synthesis by NTP sensing and other mechanisms responsible for rRNA transcription. However, it is not sufficient to determine these mechanisms in isolation. We now propose to begin to determine the relationship(s) between the multiple mechanisms affecting regulation of the synthesis of the translation apparatus during steady state growth, upshifts, and in stationary phase.